Blue EaLAMP—a specific and sensitive method for visual detection of genomic Erwinia amylovora DNA

摘要

Fire blight caused by the gram-negative bacterium Erwinia amylovora is a major disease of pome fruit like apple (Malus domestica) or pear (Pyrus communis). Detection of the pathogen is hampered by low titres usually being present during initial flower infection and the brief time frame for analysis to decide upon subsequent countermeasures. Here we describe a loop-mediated isothermal amplification of DNA (LAMP) assay for genomic DNA of Erwinia amylovora, which relies on a highly specific primer design that excludes amplification from typical DNA sources of the orchard biological system and from sample handling. The assay enables the fast detection of down to approximately 20 cfu of pure Erwinia amylovora or 100 fg genomic DNA (corresponding to approximately 25 E. amyolovora equivalents) per reaction within 45 min. Fast and reliable detection of E. amylovora in orchard samples by naked eye is achieved through a visual colour change indication with hydroxynaphthol blue. The assay avoids use of complex technical devices and is thus suitable for field testing.